Colorectal cancer (CRC) is the second most frequent cause of cancer-related death in the United States. When detected early, CRC is readily curable with surgery and in some cases with the addition of chemotherapy. However, approximately 25% of CRC patients are diagnosed with incurable metastatic disease and their predicted 5-year survival is less than 10%. Thus, novel approaches are urgently needed to improve clinical outcomes of CRC patients. Several recent studies have shown that the immunoproteasome, an alternative form of the constitutive proteasome, plays a role in a number of diseases including cancer and inflammatory diseases. In our preliminary studies, we found that LMP2, a major catalytic subunit of the immunoproteasome, is highly expressed in primary CRC tissues and cell lines compared to nonmalignant colorectal tissues. In addition, the treatment of CRC cells with 'UK-101', a small molecule that selectively inhibits LMP2, induces cell death. Interestingly, the human LMP2 has naturally occurring genetic variations including a common LMP2 polymorphism (p.60R>H, allelic frequency of 17~35%) which was previously associated with decreased proteasomal activity. However, the functional impact of LMP2 polymorphisms in CRC has never been investigated. In this proposal, we will assess the potential of the immunoproteasome-targeting approach as a novel CRC therapy using UK-101. Our hypothesis is that the antitumor effect of UK-101 is mediated by LMP2 inhibition and the therapeutic efficacy of UK-101 may be influenced by the expression level and genetic variations of LMP2 in CRC. The following specific aims are proposed;1) To determine the frequency of LMP2 expression in CRC tissues and investigate the molecular mechanisms of UK-101-induced CRC cell death, 2) To determine the impact of LMP2 polymorphisms on the proteasomal activity and sensitivity of CRC to UK-101. We expect that the results from the proposed studies will provide crucial information in evaluating the therapeutic potential of the immunoproteasome as a novel target in CRC therapy and identifying genetic and molecular factors affecting the tumor response to the immunoproteasome-targeting approach. The proposed study is the first one to explore the immunoproteasome-targeting approach in solid tumor therapy. The results from the proposed study will provide crucial data for further investigations of the in vitro and in vivo efficacy of the immunoproteasome-targeting approaches in treating CRC. PUBLIC HEALTH RELEVANCE: Although the effectiveness of bortezomib, a broadly acting proteasome inhibitor, in multiple myeloma has validated the proteasome as a cancer drug target, the broad application of bortezomib is limited due to its side effects and development of resistance. The immunoproteasome may be an alternative target in cancer therapy. Using an immunoproteasome-specific inhibitor developed in these laboratories, we will validate the immunoproteasome as a potential target for colorectal cancer treatment.